Multicellularity, stem cells, and the neoblasts of the planarian Schmidtea mediterranea

All multicellular organisms depend on stem cells for their survival and perpetuation. Their central role in reproductive, embryonic, and post-embryonic processes, combined with their wide phylogenetic distribution in both the plant and animal kingdoms intimates that the emergence of stem cells may have been a prerequisite in the evolution of multicellular organisms. We present an evolutionary perspective on stem cells and extend this view to ascertain the value of current comparative studies on various invertebrate and vertebrate somatic and germ line stem cells. We suggest that somatic stem cells may be ancestral, with germ line stem cells being derived later in the evolution of multicellular organisms. We also propose that current studies of stem cell biology are likely to benefit from studying the somatic stem cells of simple metazoans. Here, we present the merits of neoblasts, a largely unexplored, yet experimentally accessible population of stem cells found in the planarian Schmidtea mediterranea. We introduce what we know about the neoblasts, and posit some of the questions that will need to be addressed in order to better resolve the relationship between planarian somatic stem cells and those found in other organisms, including humans.     

A critical review of the role of M2PYK in the Warburg effect

It is becoming generally accepted in recent literature that the Warburg effect in cancer depends on inhibition of M₂PYK, the pyruvate kinase isozyme most commonly expressed in tumors. We remain skeptical. There continues to be a general lack of solid experimental evidence for the underlying idea that a bottle neck in aerobic glycolysis at the level of M₂PYK results in an expanded pool of glycolytic intermediates (which are thought to serve as building blocks necessary for proliferation and growth of cancer cells). If a bottle neck at M₂PYK exists, then the remarkable increase in lactate production by cancer cells is a paradox, particularly since a high percentage of the carbons of lactate originate from glucose. The finding that pyruvate kinase activity is invariantly increased rather than decreased in cancer undermines the logic of the M₂PYK bottle neck, but is consistent with high lactate production. The “inactive” state of M₂PYK in cancer is often described as a dimer (with reduced substrate affinity) that has dissociated from an active tetramer of M₂PYK. Although M₂PYK clearly dissociates easier than other isozymes of pyruvate kinase, it is not clear that dissociation of the tetramer occurs in vivo when ligands are present that promote tetramer formation. Furthermore, it is also not clear whether the dissociated dimer retains any activity at all. A number of non-canonical functions for M₂PYK have been proposed, all of which can be challenged by the finding that not all cancer cell types are dependent on M₂PYK expression. Additional in-depth studies of the Warburg effect and specifically of the possible regulatory role of M₂PYK in the Warburg effect are needed.     

Developmental mechanisms and adult stem cells for therapeutic lung regeneration

Chronic degenerative lung diseases are essentially untreatable pathological conditions. By contrast, the healthy lung has numerous mechanisms that allow for rapid repair and restoration of function following minor acute injuries. We discuss the normal endogenous processes of lung development, homeostatic maintenance and repair and consider the research strategies required for the development of methods for human therapeutic lung regeneration.     

Differentially expressed genes: OCT-4, SOX2, STAT3, CDH1 and CDH2, in cultured mesenchymal stem cells challenged with serum of women with endometriosis

Endometriosis is a common chronic gynecological disorder defined as the presence of ectopic functional endometrial tissues, outside uterine cavity, primarily on the pelvic peritoneum and the ovaries. Several studies revealed a correlation between aberrant stem-cell activity in the endometrium and endometriosis. Yet the molecular and cellular behaviors of mesnchymal stem cells in development of endometriosis are hampered by lack of invitro experiments. Our aim was to explore morphological and molecular changes associated with mesenchymal stem cells (MSCs) exposition to serum derived from women with severe endometriosis. Two cell cultures of MSCs isolated from endometrial tissues of two endometriosis-free women. Each cell culture was treated individually with the serum of women with endometriosis (experimental group/n =?7), and serum of women without endometriosis (control group/ n = 4) for 14?days. Quantitative Real-Time PCR was performed later to reveal expression of OCT-4, CDH1 and CDH2, STAT3 and SOX2 genes. Morphologically, cells showed no significant changes. However from molecular point of view, we found increased expression in OCT-4, CDH1 and CDH2. For STAT3 and SOX2 we did not find a significant difference. This study shows that endometriosis serum induced molecular changes in human endometrial MSCs (EnMSCs) that might be related to altered cell behavior which may be a step in differentiation that may be completed invivo by other factors to complete the process of transition. Further researches are needed for optimization to reach differentiation.     

Synthesis and urease inhibitory potential of benzophenone sulfonamide hybrid in vitro and in silico

This study deals with the synthesis of benzophenone sulfonamides hybrids (1–31) and screening against urease enzyme in vitro. Studies showed that several synthetic compounds were found to have good urease enzyme inhibitory activity. Compounds 1 (N′-((4′-hydroxyphenyl)(phenyl)methylene)-4′′-nitrobenzenesulfonohydrazide), 2 (N′-((4′-hydroxyphenyl)(phenyl)methylene)-3′′-nitrobenzenesulfonohydrazide), 3 (N′-((4′-hydroxyphenyl)(phenyl)methylene)-4′′-methoxybenzenesulfonohydrazide), 4 (3′′,5′′-dichloro-2′′-hydroxy-N′-((4′-hydroxyphenyl)(phenyl)methylene)benzenesulfonohydrazide), 6 (2′′,4′′-dichloro-N′-((4′-hydroxyphenyl)(phenyl)methylene)benzenesulfonohydrazide), 8 (5-(dimethylamino)-N′-((4-hydroxyphenyl)(phenyl)methylene)naphthalene-1-sulfono hydrazide), 10 (2′′-chloro-N′-((4′-hydroxyphenyl)(phenyl)methylene)benzenesulfonohydrazide), 12 (N′-((4′-hydroxyphenyl)(phenyl)methylene)benzenesulfonohydrazide) have found to be potently active having an IC₅₀ value in the range of 3.90–17.99?µM. These compounds showed superior activity than standard acetohydroxamic acid (IC₅₀?=?29.20?±?1.01?µM). Moreover, in silico studies on most active compounds were also performed to understand the binding interaction of most active compounds with active sites of urease enzyme. Structures of all the synthetic compounds were elucidated by ¹H NMR, ¹³C NMR, EI-MS and FAB-MS spectroscopic techniques.     

Hepatic lipid metabolism: effect of spermine, albumin, and Z protein on microsomal lipid formation.

Effects of spermine, bovine serum albumin, and Z protein on microsomal lipid formation from sn-glycerol 3-phosphate and [¹⁴C]palmitoyl CoA were investigated. In the presence of these agents, microsomal lipid formation was stimulated. This was attributed to the activation of sn-glycerol 3-phosphate acyltransferase and to the inhibition of palmitoyl CoA hydrolase. In addition to palmitoyl CoA, spermine also reacted with microsomal membranes in causing their aggregation, and ATP reversed the effect of spermine. Further studies indicated that the interaction of spermine with palmitoyl CoA, rather than with microsomal membranes, was responsible for the activation of glycerolipid formation or to the inhibition of palmitoyl CoA reductase. Examination of the intravesicular distribution of sn-glycerol 3-phosphate acyltransferase and palmitoyl CoA hydrolase and the effects of structural integrity of microsomal vesicles on these two membrane-bound enzymes indicated that the activation of glycerolipid formation and the inhibition of palmitoyl CoA hydrolase by spermine, bovine serum albumin, or Z protein may be closely linked with the structural integrity of microsomal vesicles.     

Pseudotyped baculovirus is an effective gene expression tool for studying molecular function during axolotl limb regeneration

Axolotls can regenerate complex structures through recruitment and remodeling of cells within mature tissues. Accessing the underlying mechanisms at a molecular resolution is crucial to understand how injury triggers regeneration and how it proceeds. However, gene transformation in adult tissues can be challenging. Here we characterize the use of pseudotyped baculovirus (BV) as an effective gene transfer method both for cells within mature limb tissue and within the blastema. These cells remain competent to participate in regeneration after transduction. We further characterize the effectiveness of BV for gene overexpression studies by overexpressing Shh in the blastema, which yields a high penetrance of classic polydactyly phenotypes. Overall, our work establishes BV as a powerful tool to access gene function in axolotl limb regeneration.     

Correlation of micro vessel density and c-Myc expression in breast tumor of mice following mesenchymal stem cell therapy

Stem cell therapy for degenerative diseases has been established; however there are controversies over the treatment of solid tumors with stem cell transplantation. In the present study, the anti-tumor action of mesenchymal stem cells (MSCs) has been examined in a mouse model of breast cancer with emphasize on tumor growth, angiogenesis and c-Myc expression in breast tumors. For this purpose, MSCs were isolated from bone marrow of Balb/c mice and characterized. A Balb/c mouse model of breast cancer was developed and subjected to cell therapy intra venous (I.V) or intra tumor (I.T) with MSCs. Tumor growth was measured using a digital caliber for until the end of experiment (30 days). Then the mice were sacrificed and their tumors were removed and processed for histopathological examination, immunohistochemical assay of CD31 and measuring of c-Myc expression using quantitative PCR. Detection of the labeled-MSCs in tumors following injection of the cells (I.V or I.T) clearly showed the homing of MSCs into tumors. Tumor growth in case of MSC-treated mice by I.V and I.T routes was inhibited by approximately 28% and 34% respectively compared to controls. The suppression of angiogenesis was reflected in Micro Vessel Density (MVD) following I.V or I.T delivery of the MSCs. c-Myc gene expression in tumor tissues of mice treated I.V or IT with MSCs was down-regulated to 28.0% and 16.0% respectively compare to control groups. In conclusion, growth inhibition of breast tumors in mice due to MSC therapy is associated with modulation of c-Myc activation and angiogenesis markers.     

Ratiometric fluorescence imaging of nuclear pH in living cells using Hoechst-tagged fluorescein

Small-molecule fluorescent sensors that allow specific measurement of nuclear pH in living cells will be valuable for biological research. Here we report that Hoechst-tagged fluorescein (hoeFL), which we previously developed as a green fluorescent DNA-staining probe, can be used for this purpose. Upon excitation at 405 nm, the hoeFL–DNA complex displayed two fluorescence bands around 460 nm and 520 nm corresponding to the Hoechst and fluorescein fluorescence, respectively. When pH was changed from 8.3 to 5.5, the fluorescence intensity ratio (F₅₂₀/F₄₆₀) significantly decreased, which allowed reliable pH measurement. Moreover, because hoeFL binds specifically to the genomic DNA in cells, it was applicable to visualize the intranuclear pH of nigericin-treated and intact living human cells by ratiometric fluorescence imaging.     

Isoxazolo[3,4-d]pyridazinones positively modulate the metabotropic glutamate subtypes 2 and 4

Isoxazolo[3,4-d] pyridazinones ([3,4-d]s) are selective positive modulators of the metabotropic glutamate receptors (mGluRs) subtypes 2 and 4, with no functional cross reactivity at mGluR_1a, mGLuR₅ or mGluR₈. Modest binding for two of the [3,4-d]s is observed at the allosteric fenobam mGluR₅ site, but not sufficient to translate into a functional effect. The structure activity relationship (SAR) for mGluR₂ and mGluR₄ are distinct: the compounds which select for mGluR₂ all contain fluorine on the N-6 aryl group. Furthermore, the [3,4-d]s in this study showed no significant binding at inhibitory GABA_A, nor excitatory NMDA receptors, and previously we had disclosed that they lack significant activity at the System Xc-Antiporter. A homology model based on Conn’s mGluR₁ crystal structure was examined, and suggested explanations for a preference for allosteric over orthosteric binding, subtype selectivity, and suggested avenues for optimization of efficacy as a reasonable working hypothesis.     

Genomic,molecular evolution,and expression analysis of NOX genes in soybean (Glycine max)

Reactive oxygen species (ROS) are versatile signaling molecules in sensing stresses and play critical roles in signaling and development. Plasma membrane NADPH oxidases (NOXs) are key producers of ROS, and play important roles in the regulation of plant-pathogen interactions. Here, we performed a comprehensive analysis of the NOX gene family in the soybean genome (Glycine max) and 17 NOX (GmNOX) genes were identified. Structural analysis revealed that the GmNOX proteins in soybean were as conserved as those in other plants. 8 duplicated gene pairs were formed by a Glycine-specific whole-genome duplication (WGD) event approximately 13 million years ago (Mya). The Ka/Ks ratios of GmNOX genes ranged from 0.04 to 0.28, suggesting that the GmNOX family had undergone purifying selection in soybean. Gene expression patterns showed different expression of these duplicate genes, suggesting that the GmNOXs were retained by substantial subfunctionalization during the soybean evolutionary processes. Subsequently, the expression of GmNOXs in response to drought and phytohormones were characterized via qPCR. Importantly, four GmNOXs showed strong expression in nodules, pointing to their probable involvement in nodulation. Thus, our results shed light on the evolutionary history of this family in soybean and contribute to the functional characterization of GmNOX genes in soybean.     

Molecular phylogeny of the extinct Steller's sea cow and other Sirenia species based on their complete mitochondrial genomes

The Steller's sea cow – Hydrodamalis gigas (Dugongidae: Sirenia) – is an extinct herbivorous marine mammal which inhabited the North Pacific Ocean during the Pleistocene and Holocene. H. gigas was the largest member of the Sirenia order and disappeared in the middle of the 18th century. Here, we present the complete sequence of the mitochondrial genome of this extinct animal. The Steller's sea cow mitochondrial DNA (mtDNA) is 16,872 base pairs (bp) in length and contains a set of mitochondrial genes typical for mammals. Phylogenetic analysis based on complete mitochondrial genomes of the sirenian species allows accurate assessment of the degree of their mitogenomic diversification during millions of years of evolution.     

Synthesis and antiproliferative activity of derivatives of the phyllanthusmin class of arylnaphthalene lignan lactones

A series of arylnaphthalene lignan lactones based on the structure of the phyllanthusmins, a class of potent natural products possessing diphyllin as the aglycone, has been synthesized and screened for activity against multiple cancer cell lines. SAR exploration was performed on both the carbohydrate and lactone moieties of this structural class. These studies have revealed the importance of functionalization of the carbohydrate hydroxy groups with both acetylated and methylated analogues showing increased potency relative to those with unsubstituted sugar moieties. In addition, the requirement for the presence and position of the C-ring lactone has been demonstrated through reduction and selective re-oxidation of the lactone ring. The most potent compound in this study displayed an IC₅₀ value of 18?nM in an HT-29 assay with several others ranging from 50 to 200?nM. In an effort to elucidate their potential mechanism(s) of action, the DNA topoisomerase IIa inhibitory activity of the most potent compounds was examined based on previous reports of structurally similar compounds, but does not appear to contribute significantly to their antiproliferative effects.     

Phylogenetic analyses of Bradyrhizobium symbionts associated with invasive Crotalaria zanzibarica and its coexisting legumes in Taiwan

In this study, the genetic diversity and identification of Bradyrhizobium symbionts of Crotalaria zanzibarica, the most widely-distributed invasive legume in Taiwan, and other sympatric legume species growing along riverbanks of Taiwan were evaluated for the first time. In total, 59 and 54 Bradyrhizobium isolates were obtained from C. zanzibarica and its coexisting legume species, respectively. Based on the multilocus sequence analysis (MLSA) of concatenated four housekeeping genes (dnaK-glnII-recA-rpoB gene sequences, 1901 bp), the 113 isolates displayed 53 unique haplotypes and grouped into 21 clades. Of these clades, 11 were found to be congruent to already defined Bradyrhizobium species, while the other 10 clades were found to not be congruent to any defined species. In particular, the C. zanzibarica isolates belong to 14 MLSA clades, six of which overlapped with the isolates of coexisting legumes. According to the nodA gene sequences (555 bp) obtained from the 105 isolates, these isolates were classified into three known nodA clades, III.2, III.3 and VII and were further clustered into 10 groups. Furthermore, the C. zanzibarica isolates were clustered into 8 nodA groups, five of which overlapped with the isolates from coexisting legumes. Additionally, the nodA genes of the isolates from native species were dominated by Asian origin, while those from C. zanzibarica were dominated by American origin. In conclusion, C. zanzibarica is a promiscuous host capable of recruiting diverse Bradyrhizobium symbionts, some of which are phylogenetically similar to the symbionts of coexisting legumes in Taiwan.     

Molecular dynamics simulations of the chemokine CCL2 in complex with pull down-derived heparan sulfate hexasaccharides

Background

Binding of chemokines to glycosaminoglycans (GAGs) is a crucial step in leukocyte recruitment to inflamed tissues.